Noradrenaline Overflow Research Articles

Corelease of noradrenaline and ATP by brief pulse trains in guinea-pig vas deferens.

Contractions and overflow of tritium and ATP elicited by single electrical pulses or short pulse trains were studied in the guinea-pig isolated vas deferens preincubated with [3H]-noradrenaline. ATP was measured using the luciferase technique. A single pulse caused only a small contraction and minimal tritium and ATP overflow. In contrast, trains of 6 pulses elicited marked contractions as well as tritium and ATP overflow. In experiments with 6 pulses/100 Hz, prazosin 0.3 microM reduced the contraction by 73%, did not change the evoked overflow of tritium, and reduced the evoked overflow of ATP by 85%. Suramin 300 microM reduced the contraction by 69% but changed neither the evoked overflow of tritium nor that of ATP. The combination of prazosin 0.3 microM and suramin 300 microM abolished the contraction, did not change the evoked overflow of tritium, and reduced the evoked overflow of ATP by 70%. When 6 pulses were applied at frequencies of 1, 2, 10 or 100 Hz, all responses increased with frequency up to a maximum at 10 Hz, but contractions and the evoked overflow of ATP increased with frequency to a greater extent than the evoked overflow of tritium. A similar frequency overflow relationship was observed when the medium contained prazosin 0.3 microM and suramin 300 microM (and evoked ATP overflow was greatly reduced). Yohimbine 1 microM did not affect the overflow of tritium evoked by 6 pulses/100 Hz but increased that evoked by 6 pulses/10 Hz. The results demonstrate an overflow of both noradrenaline and ATP in response to short pulse trains.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibitory effects of nicorandil on sympathetic coronary vasoconstriction

The aim was to investigate whether nicorandil suppresses the rise in coronary vascular resistance that occurs during stimulation of the sympathetic nerve supply to the heart and, if so, what are the mechanisms of action. The effects of nicorandil on coronary vascular resistance during ventrolateral cardiac nerve stimulation and on the reactivity of the coronary vasculature to intracoronary infusion of noradrenaline or neuropeptide Y (NPY) were examined under beta receptor blockade. The effects of nicorandil on the overflow of noradrenaline and NPY during ansae subclaviae stimulation were compared with those in a control group and in a group treated with glyceryl trinitrate and N omega-nitro-L-arginine (L-NNA) under both alpha and beta receptor blockade with vagotomy. Intracoronary infusion of nicorandil decreased coronary vascular resistance prior to cardiac nerve stimulation, and during stimulation it suppressed the percentage increase in resistance from the prestimulation value. Nicorandil suppressed the reactivity of the coronary vasculature to exogenous noradrenaline and NPY. Intra-atrial infusion of nicorandil significantly reduced the overflow of NPY but not of noradrenaline during stimulation of the ansae subclaviae at 20 Hz. This suppressive effect was not observed in the glyceryl trinitrate + L-NNA group. Nicorandil reduces sympathetic coronary vasoconstriction by decreasing the reactivity of the vasculature to sympathetic neurotransmitters and by suppressing NPY overflow during cardiac sympathetic nerve stimulation. The suppressive action on NPY overflow is thought to be due to the opening of ATP sensitive potassium channels in the sympathetic nerve endings rather than to a glyceryl trinitrate-like action of nicorandil.

Norepinephrine and dihydroxyphenylglycol effluxes from sympathetic nerve endings during hypoxia and reoxygenation in the isolated rat heart.

The present experiments were carried out in isolated rat hearts perfused according to the Langendorff method at a constant pressure of 10 kPa. The aim was to measure norepinephrine (NE) overflow and its deaminated metabolite dihydroxyphenylglycol (DOPEG) by changing the composition of the buffer perfusing the heart to simulate hypoxia. When aerobic and glycolytic pathways were simultaneously reduced, NE and DOPEG overflow increased 711 and 145%, respectively, after 30 min, compared with control values of 0.45 +/- 0.06 and 0.66 +/- 0.7 ng.min-1.g-1 of heart (n = 8, p < 0.05). Whereas NE leakage decreased sharply after reoxygenation and glucose addition, DOPEG continued to increase up to 260% after 5 min of normal reperfusion. This mechanism was calcium independent and inhibited by 80% with desipramine (1 microM), confirming the role of the uptake I carrier, which reversed its normal transport direction. Neuropeptide Y, a marker of exocytotic release, did not increase in the perfusate with the progression of hypoxia, which supports the hypothesis of a nonexocytotic release. Tyramine (1 microM) significantly enhanced NE outflow by displacing the amine from its storage vesicles through a calcium-independent mechanism, indicating that a pool of NE was still available. In the presence of 1 microM clorgyline (a monoamine oxidase A inhibitor) but not deprenyl (a monoamine oxidase B inhibitor), NE outflow increased 934% and DOPEG only 40% at 30 min (n = 6, p < 0.05 versus control hearts).(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of neonatal exposure to clonidine on basal and activated central noradrenaline metabolism and in vivo overflow

The occurrence of persistent effects of chronic neonatal exposure to the α 2- adrenoceptor agonist clonidine was investigated by determination of tissue concentrations of monoamines and metabolites and in vivo overflow of noradrenaline and its metabolites, in various rat brain regions during adulthood. Rat pups were treated with clonidine from postnatal day 10–20 and all measurements were carried out between postnatal day 40 and 58. Tissue concentrations of monoamines and metabolites of the early clonidine-treated rats did not differ significantly from the control group. A challenge with yohimbine did not reveal altered responses of monoaminergic systems, except for the failure of an increased serotonergic activity in the medulla pons. In vivo microdialysis measurements revealed an elevated basal extracellular noradrenaline level in amygdala, but not in frontal cortex and hippocampus. Pharmacological challenge in vivo with idazoxan did not reveal differences between clonidine- and saline-exposed rats. These results confirm previous findings that continuous activation of α 2- adrenoceptors during a particular period of rat brain development may result in long-lasting but small changes in monoaminergic activity. However, these alterations are not very consistent and may depend on the parameter chosen to reflect monoaminergic activity and are not revealed more clearly by activating (challenging) the noradrenaline system with α 2- adrenoceptor antagonists.

Pre- and postganglionic stimulation-induced noradrenaline overflow is markedly facilitated by a prejunctional β2-adrenoceptor-mediated control mechanism in the pithed rat

The aim of the study was to further explore the prejunctional beta-adrenoceptor-mediated control mechanism of noradrenaline release from sympathetic nerves in response to preganglionic nerve stimulation (PNS) and local nerve stimulation of the portal vein, respectively, in the pithed rat. Baseline values as well as the increments of mean arterial blood pressure (delta-BP), heart rate (delta-HR) and plasma noradrenaline levels (delta-NA) in response to four PNS episodes (0.8 Hz, 3 ms, 75 V for 45 s at 20 min intervals), respectively, were evaluated. Fenoterol administration (0.25 mg/kg, i.v.) reduced significantly the basal blood pressure but did not alter delta-BP in response to PNS. Basal heart rate markedly increased after fenoterol without any further change in heart rate induced by PNS. The beta 1-selective antagonist CGP 20712A attenuated delta-BP in response to PNS and prevented the fenoterol-induced increase in basal heart rate. The beta 2-selective antagonist ICI 118,551 per se did not change the blood pressure and heart rate values, but antagonized the fenoterol-induced decrease in basal blood pressure. Fenoterol enhanced plasma delta-NA in response to PNS by 105% in comparison to the corresponding control value. This effect of fenoterol could be blocked by pretreatment with ICI 118,551 but not with CGP 20712A (a selective beta 1-adrenoceptor antagonist) which per se did not significantly change plasma delta-NA. Repeated local stimulation of the portal vein (S1-S3, 2 Hz, 3 ms, 10 mA, for 120 s at 30 min intervals) increased portal plasma noradrenaline without changing mean blood pressure and heart rate in pithed rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Increasedα 2-adrenoceptor mediated regulation of adult rat brain noradrenaline overflow after chronic neonatal exposure to propranolol; a microdialysis study

Direct and persistent effects of chronic neonatal administration of the β-adrenoceptor antagonist propranolol on brain noradrenergic activity were investigated by measuring tissue concentrations of noradrenaline and its metabolites and in vivo overflow of noradrenaline during adulthood. Rat pups were chronically treated with propranolol from postnatal day 1 to day 10. Determination of monoamine metabolism after the last injection showed an increase in noradrenaline metabolism in frontal cortex, limbic system and hippocampus of propranolol-exposed rats, but 47 days after this last injection it was apparent that these effects were not long-lasting. Moreover, basal noradrenaline overflow in vivo in the hippocampus of 40–55 day-old propranolol-exposed rats did not differ from that in controls. However, the regulation of noradrenaline release seemed to have been altered, since a pharmacological challenge with theα 2-adrenoceptor antagonist idazoxan induced an enhanced increase in the in vivo noradrenaline overflow in propranolol-exposed rats compared to controls. It is suggested that the neonatal β-blockade induced a supersensitivity of the presynapticα 2-adrenoceptor. The precise mechanism underlying this effect has to be elucidated.

Effects of human recombinant interleukins on stimulation-evoked noradrenaline overflow from the rat perfused spleen.

Experiments were carried out in the isolated spleen of the rat to study in a lymphoid organ the influence of interleukins (ILs) on noradrenaline release. Spleens were perfused with Tyrode's solution and the overflow of endogenous noradrenaline was determined by HPLC with electrochemical detection. Perivascular electrical stimulation (4 or 10 Hz, 20-28 mA, 2 min) caused an increase in noradrenaline overflow and in perfusion pressure, both of which were markedly reduced by perfusion with Ca(2+)-free solution, abolished by tetrodotoxin, unaffected by hexamethonium, and subject to alpha 2-adrenoceptor- and muscarinic receptor-mediated modulation as shown by the effects of rauwolscine and methacholine. Human recombinant IL-1 beta and IL-2 and mouse recombinant IL-2 10 ng/ml failed to affect the evoked overflow of noradrenaline after an exposure time of 15 min. In contrast, human recombinant IL-1 beta and IL-2 0.1 ng/ml reduced the evoked overflow after exposure for 80 min; the inhibition tended to increase 30 min later despite washout. Murine recombinant IL-2 1.2 ng/ml caused no change after contact with the tissue for 80 min but there was an inhibition 30 min later after washout. Human recombinant IL-6 (0.1 ng/ml) caused no significant change. The inhibitory effect of low concentrations of IL-1 beta and IL-2 supports the idea that locally produced mediators of the immune system may affect neuronal function.

Focal and systemic cocaine differentially affect extracellular norepinephrine in the locus coeruleus, frontal cortex and hippocampus of the anaesthetized rat

The purpose of this study was to characterize and compare the effects of cocaine on norepinephrine (NE) overflow in the forebrain and somatodendritic regions of anaesthetized rats with microdialysis. Intraperitoneal injections of cocaine (20 mg/kg) failed to increase NE overflow in the hippocampus and the frontal cortex but did elevate NE in the region of the locus coeruleus. Focal application of cocaine (1–100 μM) via the dialysis probe into the region of the locus coeruleus also produced a concentration dependent elevation of extracellular NE. In the terminal regions the application of focal cocaine (1–100 μM) showed a differential effect, with a concentration dependent increase in extracellular NE in the hippocampus, whilst in the frontal cortex only the highest concentration of cocaine (100 μM) elevated extracellular NE. The regional differences seen following focal applications in this study may be related to differences in transporter function in the three brain areas or to differences in the affinity for cocaine. The inability of systemically administered cocaine to increase hippocampal and cortical NE is probably related to its predominant actions in the somatodendritic region.

Functional relation between nitric oxide and noradrenaline for the modulation of vascular tone in rat mesenteric vasculature

As previously reported, N omega-nitro-L-arginine (L-NNA), an inhibitor of nitric oxide (NO) synthesis, decreased transmural field stimulation (TFS)-induced noradrenaline overflow from the isolated perfused rat mesenteric vasculature attached to the intestine. The decrease was attenuated by L-arginine. This suggests that NO may increase noradrenaline release (Yamamoto et al. 1993). The present experiments with this preparation were done in order to monitor changes in vascular perfusion pressure caused by TFS or by noradrenaline infusion in parallel with those in the noradrenaline outflow caused by TFS in the presence of atropine (0.1 mumol/l) (to block acetylcholine-induced release of endothelial NO) and of indomethacin (3 mumol/l) (to inhibit L-NNA-induced production of vasoconstrictor prostanoids). (1) TFS (2-10 Hz) caused a frequency-dependent increase in noradrenaline overflow and perfusion pressure. (2) L-NNA (10 and 30 mumol/l) caused a concentration-dependent inhibition of TFS-induced noradrenaline overflow, whereas the TFS-induced pressure increase was augmented by L-NNA in a concentration-dependent manner. At any given concentration of L-NNA, the potentiation of vasoconstriction by L-NNA became greater in magnitude as the frequency of the TFS was raised. (3) Infusion of noradrenaline (0.38-6 nmol) caused a dose-dependent increase in perfusion pressure up to a value comparable with that caused by TFS. The pressure increase in response to noradrenaline infusion was also enhanced by L-NNA, relatively, to a greater extent than the enhancement, by L-NNA, of the pressure response to TFS.(ABSTRACT TRUNCATED AT 250 WORDS)

Pharmacological characterization and anatomical localization of prejunctional beta-adrenoceptors in the rat kidney.

1. The subtype and anatomical localization of beta-adrenoceptors mediating facilitation of stimulus-induced overflow of noradrenaline ('prejunctional beta-adrenoceptors') are not conclusively known to date. The present study was undertaken to characterize these receptors by use of pharmacological methods as well as to define their localization (prejunctional or postjunctional) with radio-ligand binding and autoradiography techniques combined with surgical denervation of the sympathetic innervation to the rat kidney. 2. Exposure of the kidney to (-)-isoprenaline, the nonselective beta-adrenoceptor agonist, resulted in a dose-dependent facilitation of stimulus-induced neurotransmitter overflow. This response was inhibited by propranolol, the beta 1- and beta 2-adrenoceptor antagonist, with a pA2 of 9.20 suggesting that the prejunctional beta-adrenoceptors are not of the beta 3-subtype. 3. The rank order of potency and potency ratios of beta-adrenoceptor agonists at renal prejunctional beta-adrenoceptors (EC50 for agonist/EC50 for (-)-isoprenaline) were: (-)-isoprenaline (1) > procaterol (2) > salbutamol (3) > adrenaline (10) > (+)-isoprenaline (25). However, dobutamine, the beta 1-adrenoceptor agonist, failed to enhance stimulus-induced overflow of noradrenaline. These results are indicative of the presence of beta 2-adrenoceptors as prejunctional beta-adrenoceptors. 4. Facilitation elicited by (-)-isoprenaline and procaterol, the selective beta 2-adrenoceptor agonist, was inhibited by ICI 118,551, the selective beta 2-adrenoceptor antagonist, with pKb values of 9.20 and 9.35, respectively at renal prejunctional beta-adrenoceptors. Similarly, the pKb values of metoprolol, the selective beta 1-adrenoceptor antagonist, at renal prejunctional beta-adrenoceptors were determined to be 6.25 and 6.18 against (-)-isoprenaline and procaterol, respectively. These results suggest the presence of a homogeneous population of beta 2-adrenoceptors as prejunctional beta-adrenoceptors. 5. Radio-ligand binding analysis of renal beta-adrenoceptors revealed the prevalence of the beta 1-subtype as compared to the beta 2-subtype (63% vs 37%). However, surgical denervation of the rat kidney, resulting in more than 90% reduction in renal noradrenaline content, selectively reduced the beta 2-adrenoceptor population by 80%, implying the presence of beta 2-adrenoceptors on renal sympathetic nerve terminals. 6. Autoradiographic analysis demonstrated the presence of beta 1-adrenoceptors on cortical structures such as glomeruli and tubules. beta-Adrenoceptors were found to be present on tubules (minor population), collecting tubules in outer medulla and the adventitia and adventitial-medial border of intraparenchymal branches of the renal artery. Surgical denervation of the rat kidney resulted in the disappearance of Beta2-adrenoceptors associated with the intraparenchymal branches, without affecting the Beta-adrenoceptor populations at other sites. These results support the notion that the Beta2-subtype is present on renal sympathetic nerve terminals and demonstrate that these prejunctional Beta2-adrenoceptors are associated with the renal vasculature and not with renal tubules.7. The results of the present investigation demonstrate that renal prejunctional Beta-adrenoceptors are of the Beta2-subtype in nature. These receptors are present on sympathetic nerve terminals which are associated with the renal vasculature.

Characterization of the prejunctional muscarinic receptors mediating inhibition of evoked release of endogenous noradrenaline in rabbit isolated vas deferens

The aim of the present study was to characterize the prejunctional modulation of evoked release of endogenous noradrenaline in rabbit vas deferens by the use of muscarinic receptor agonists and subtype-preferring antagonists. Vasa deferentia of the rabbit were stimulated electrically by trains of 120 pulses delivered at 4 Hz or trains of 30 pulses at 1 Hz. The inhibition by muscarinic agonists of the stimulation-evoked overflow of endogenous noradrenaline in the absence and presence of antagonists was used to determine affinity constants for antagonists. These values were compared with those observed at putative M1 receptors inhibiting neurogenic twitch contractions in the rabbit vas deferens and with affinity data obtained at M1(m1)-M4(m5) receptors in functional studies and binding experiments. The evoked overflow of noradrenaline from sympathetic nerves was enhanced by the A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), the P2 purinoceptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) and indomethacin, indicating a tonic inhibition by endogenous A1 and P2 purinoceptor agonists and prostanoids, respectively. The stimulation-evoked overflow at 4 Hz was not sensitive to inhibition by the muscarinic agonists methacholine or 4-(4-chlorophenylcarbamoyloxy)-2-butynyltrimethylammonium iodide (4-Cl-McN-A-343). In contrast, at a stimulation frequency of 1 Hz the evoked noradrenaline release was decreased by muscarinic agonists (EC50): arecaidine propargyl ester (0.062 microM), 4-Cl-McN-A-343 (0.32 microM), 4-(4-fluorophenylcarbamoyloxy)-2-butynyl-N-methyl-pyrrolidinium tosylate (4-F-PyMcN+; 0.48 microM) and methacholine (0.86 microM). The affinity constants of most of the muscarinic antagonists [atropine: pKB = 9.47; (R)-trihexyphenidyl: pKB = 9.18; pirenzepine: pA2 = 7.68; methoctramine: pKB = 6.90] are consistent with estimates of these antagonists at M1(m1) receptors determined in various functional and binding studies. The high antagonistic potency of pirenzepine and (R)-trihexyphenidyl and the agonistic activity of 4-F-PyMcN+ argue for the involvement of M1, and against that of M2 and M3 receptors in the inhibition of evoked noradrenaline overflow. However, the high apparent pKB of 8.30 for himbacine is not in accordance with an M1 receptor; by contrast, it would be compatible with the presence of M2 or M4 receptors. The potencies of the tested muscarinic agonists and antagonists largely agree with those obtained for the inhibition of neurogenic twitch responses (0.05 Hz) in the rabbit vas deferens.(ABSTRACT TRUNCATED AT 400 WORDS)

Regional differences in the in vivo regulation of the extracellular levels of noradrenaline and its metabolites in rat brain

Microdialysis was used to determine extracellular levels of both noradrenaline and its metabolites in several brain regions of rats under basal conditions and in response to drugs selective for theα 2-adrenoceptor to study regional differences in the regulation of noradrenaline overflow. Basal overflow of noradrenaline was about 1.3 fmol/min in frontoparietal cortex, amygdala and hippocampus and in the medial prefrontal cortex 2.4 fmol/min was measured, whereas the overflow of the noradrenaline metabolites 3,4-dihydroxyphenylglycol and 3-methoxy-4-hydroxyphenylglycol was 10-fold higher. After correction for recovery and membrane length no regional differences in the basal overflow of noradrenaline (NA) were found. There were, however, regional differences in the drug-induced effects: locally applied moxonidine decreased extracellular noradrenaline stronger in the frontoparietal cortex than in the medial prefrontal cortex. The increase in noradrenaline overflow caused by idazoxan (10 −4M) was stronger in frontoparietal cortex than in amygdala and hippocampus. The metabolites were also generally decreased by moxonidine and increased by idazoxan, although less markedly. The present study shows that the regulation of noradrenaline overflow by the presynapticα 2-autoreceptor was stronger in cortical regions than in amygdala and hippocampus. In those latter regions the uptake mechanisms probably plays a relatively more important role in the regulation of noradrenaline overflow.

Dual effect of nicotine on cardiac noradrenaline release during metabolic blockade.

Nicotine-induced noradrenaline was investigated in perfused guinea pig hearts subjected to metabolic blockade that was caused either by anoxia or by cyanide intoxication. Noradrenaline, neuropeptide Y, and dihydroxyphenylethyleneglycol (DOPEG) were determined in the coronary venous overflow Neuropeptide Y is a sympathetic cotransmitter of noradrenaline, and concomitant release of both transmitters indicates an exocytotic, calcium-dependent release mechanism, whereas neuropeptide Y overflow does not occur during nonexocytotic noradrenaline release. Nonexocytotic, calcium-independent noradrenaline release, however, is associated with an increase of DOPEG overflow, which is the main intraneuronal metabolite of noradrenaline formed by monoamine oxidase if oxygen is present. Anoxia per se caused a nonexocytotic release of noradrenaline starting after 10 min of anoxia and reaching peak levels at 30 min. During anoxia, nicotine (3 and 10 mumol/l) accelerated and enhanced noradrenaline overflow, i.e., the period between the onset of anoxia and the begin of noradrenaline release was shortened and peak levels were increased. Nicotine-induced noradrenaline release was accompanied by neuropeptide Y overflow. The action of nicotine was further evaluated during energy depletion caused by cyanide. As anoxia did, cyanide administration alone resulted in noradrenaline release. In accordance with a nonexocytotic mechanism and due to the presence of oxygen, this release of noradrenaline was accompanied by an increase of DOPEG. When added 10 min after the onset of energy depletion, nicotine (10 mumol/l) caused a brief but marked enhancement of exocytotic noradrenaline release, since this release was calcium-dependent and was accompanied by a significant rise of neuropeptide Y overflow. In absence of extracellular calcium to avoid exocytosis, concomitant administration of nicotine (3-100 mumol/l) and cyanide caused a concentration-dependent acceleration of both the overflow of noradrenaline and DOPEG, whereas overflow of neuropeptide Y was not increased, thus indicating a nonexocytotic release mechanism. In conclusion, the application of nicotine during myocardial energy depletion increases overflow of noradrenaline by both calcium-dependent exocytotic release and calcium-independent nonexocytotic release mechanisms.

P-543 - Involvement of nitric oxide (NO) in cooling-induced decrease in transmural field stimulation (TFS)-evoked noradrenaline (NA) overflow in isolated rat vasculature.

P-543 - Involvement of nitric oxide (NO) in cooling-induced decrease in transmural field stimulation (TFS)-evoked noradrenaline (NA) overflow in isolated rat vasculature.

Intermittent release of noradrenaline by single pulses and release during short trains at high frequencies from sympathetic nerves in rat tail artery

As shown by electrophysiological analysis, the release of the sympathetic co-transmitter adenosine 5′-triphosphate (ATP) from individual release sites is monoquantal and intermittent; the average release probability may be as low as 0.01. 1–3 Indirect evidence from biochemical studies of noradrenaline overflow is compatible with a similar monoquantal, low probability release of noradrenaline as well. 10 In the present study our first aim was to address this issue more directly in rat tail artery, using continuous amperometry to monitor in real time the release of noradrenaline from a relatively small number of sympathetic nerve varicosities. The results seem to provide the first direct evidence that noradrenaline, similarly to ATP, may be released intermittently during nerve stimulation at low frequency. Our second aim was to use the same technique to study the release of noradrenaline caused by nerve stimulation with single pulses or short trains (two to eight pulses) at high frequencies. The results show that during stimulation at 20 Hz the peak amplitude of the noradrenaline oxidation current response grew linearly with the train length, but at 50 Hz the curve describing this growth was sigmoid in shape.

Mechanisms of noradrenaline release in the anoxic heart of the rat.

The aim was to examine the time course of exocytotic and "spontaneous" noradrenaline overflow and the influence of an Uptake1 inhibitor, desipramine, in rat hearts subjected to anoxic and substrate-free perfusion. Hearts were perfused with a constant flow and exocytotic noradrenaline overflow was elicited either by electrical stimulation of the left stellate ganglion or by K+ depolarisation. Noradrenaline overflow was measured by HPLC. Energy depletion for a period of 30 min resulted in an enhanced spontaneous noradrenaline overflow and a progressive decline in the nerve stimulation induced noradrenaline overflow. However, noradrenaline overflow induced by 40 mM K+ was enhanced by three- to fourfold in the energy depleted conditions. During anoxia, desipramine (0.3 microM) inhibited the spontaneous noradrenaline overflow and partly increased, in the early phase of anoxia, noradrenaline overflow by nerve stimulation, but showed no effect on K+ induced overflow. Further experiments showed that K+ at 10 mM failed to evoke noradrenaline overflow in normoxic hearts but induced a significant overflow in energy depleted hearts, either in the presence or absence of desipramine; quantities of noradrenaline overflow in response to 10-40 mM K+ were substantially higher in anoxia. This difference in noradrenaline overflow caused by K+ during normoxia and anoxia was partly narrowed by desipramine which enhanced overflow in normoxia. "Spontaneous" and exocytotic noradrenaline release coexist within the 30 min period of anoxia but their responses to Uptake1 inhibitor differ. K(+)-induced noradrenaline overflow was markedly augmented by energy depletion due to a combination of failed neuronal reuptake and enhanced exocytosis.

Clinical Application of Noradrenaline Spillover Methodology: Delineation of Regional Human Sympathetic Nervous Responses

The proportionality which in general exists between rates of sympathetic nerve firing and the overflow of noradrenaline into the venous drainage of an organ provides the experimental justification for the use of measurements of noradrenaline in plasma as a biochemical measure of sympathetic nervous function. Static measurements of noradrenaline plasma concentration have several limitations. One is the confounding influence of noradrenaline plasma clearance on plasma concentration. Other drawbacks include the distortion arising from antecubital venous sampling (this represents but one venous drainage, that of the forearm), and the inability to detect regional differentiation of sympathetic responses. Clinical regional noradrenaline spillover measurements, performed with infusions of radiolabelled noradrenaline and sampling from centrally placed catheters, and derived from regional isotope dilution, overcome these deficiencies. The strength of the methodology is that sympathetic nervous function may be studied in the internal organs not accessible to nerve recording with microneurography. Examples of the regionalization of human sympathetic responses disclosed include the preferential activation of the cardiac sympathetic outflow with mental stress, cigarette smoking, aerobic exercise, cardiac failure, coronary insufficiency, essential hypertension and in ventricular arrhythmias, and the preferential stimulation or inhibition of the renal sympathetic nerves with low salt diets and mental stress, and with exercise training, respectively. By application of the same principles, regional release of the sympathetic cotransmitters neuropeptide Y and adrenaline can be studied in humans. Cotransmitter release, however, is detected only with some difficulty. In restricted circumstances we find evidence of regional cotransmitter release to plasma, such as the release of neuropeptide Y from the heart at the very high rates of sympathetic nerve firing occurring with aerobic exercise, and cardiac adrenaline release also with exercise and after loading of the neuronal adrenaline pool by intravenous infusion of adrenaline.

Ovarial steroids and methionine-enkephalin modulation of adrenergic transmission in rabbit oviduct

1. 1. The effect of methionine-enkephalin on the [ 3H]-noradrenaline ([ 3H]-NA) overflow and on contractions evoked by field electrical stimulation (FES) and by exogenous NA were studied in vitro in the isthmic part of oviduct of rabbits (untreated and treated with estradiol or progesterone). 2. 2. The evoked tritium overflow (which reflected [ 3H]-NA overflow) was determined by liquid scintillation spectrometry. 3. 3. Field electrical stimulation of 4 Hz (trains of 40 pulses, 0.3 msec) evoked guanethidine-sensitive contractions. 4. 4. In all groups of animals methionine-enkephalin dose dependently decreased FES-evoked contractions but not those evoked by exogenous NA. 5. 5. The amount of tritium overflow evoked by 4 Hz stimulation (600 pulses, 1 msec) was significantly lower in tissues from estradiol treated (1.16 ± 0.19%) compared with those obtained in tissues from untreated (1.82 ± 0.22%) and progesterone treated (2.07 ± 0.21%) rabbits. Methionine-enkephalin, 1 μM, decreased the evoked tritium overflow in the isthmus from untreated rabbits by 36.1 ± 3.6%, in estradiol treated by 22.8 ± 2.9% and in progesterone treated by 52.3 ± 4.5%. 6. 6. The results suggest that the methionine-enkephaline effect on FES-evoked contractions could be due to a prejunctional effect on the adrenergic terminals and that there is a hormonal dependence of the opioid effect on [ 3H]-NA overflow.

The link between acute haemodynamic adrenergic beta-blockade and long-term effects in patients with heart failure. A study on diastolic function, heart rate and myocardial metabolism following intravenous metoprolol.

The present study was performed to find possible mechanisms linking the early effects of beta-blockade with the observed long-term effects in patients with heart failure. In 57 patients with heart failure, 13 +/- 3.1 mg of metoprolol was given intravenously. The patients were investigated by invasive haemodynamics (n = 34), including collection of myocardial metabolic data during atrial pacing stress (n = 16), by radionuclide angiography during physiological atrial pacing (n = 13), and by a bedside evaluation (n = 10). Diastolic function, measured by early peak filling rate, followed changes in heart rate, but was similar when heart rate was held constant by atrial pacing before and after beta-blockade. Following beta-blockade and slower heart rates, diastolic filling volumes were redistributed to late diastole. Metoprolol induced a parallel decrease in coronary sinus flow and myocardial oxygen consumption. Myocardial oxygen consumption following beta-blockade decreased both during spontaneous rhythm (25 +/- 15 to 16 +/- 8.8 ml min-1; P = 0.006), and during atrial pacing stress (30 +/- 13 to 23 +/- 11 ml.min-1; P = 0.004). Cardiac index decreased owing to reduction of heart rate (2.3 +/- 1.0 to 1.9 +/- 0.64 l.min-1.m2; P = 0.0003), while left ventricular filling pressure was unchanged. Ejection fraction and ventricular volumes were unaltered following atrial pacing or beta-blockade. There was a reflex increase in noradrenaline concentration after beta-blockade injection (0.96 +/- 0.66 to 1.20 +/- 0.91 nmol.l-1; P = 0.002), whereas myocardial noradrenaline overflow was unchanged. There was a trend towards an increase in myocardial lactate consumption after beta-blockade administration during atrial pacing stress. It is suggested that the surprisingly good tolerability seen after acute administration of beta-blockers to patients with severe heart failure may be explained by prolongation of the diastolic filling phase, which outweighs the negative inotropic effects. The reduced myocardial metabolic demand may allow the failing myocardium to recover and explain the excellent long-term effect on heart function following beta-blockade treatment.

ATP and endogenous agonists inhibit evoked [3H]-noradrenaline release in rat iris via A1 and P2y-like purinoceptors.

Effects of ATP, adenosine and purinoceptor antagonists on field stimulation-evoked (3 Hz, 2 min) [3H]-noradrenaline overflow were investigated in the rat isolated iris. ATP and adenosine inhibited the evoked overflow of [3H]-noradrenaline. 1,3-Dipropyl-8-cyclopentylxanthine (DPCPX) shifted the concentration-response curve of ATP to the right in a concentration-dependent manner, but with a potency (-log KB = 7.88) much lower than expected for an A1 adenosine receptor. In the continuous presence of DPCPX, the ATP-induced prejunctional inhibition was unaffected by suramin (100 mumol/l) and DIDS (4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid, 50 mumol/l) but was antagonized by the P2Y-receptor antagonist cibacron blue (= reactive blue 2; 30 and 100 mumol/l, -log KB = 4.7) and alpha,beta-methylene-ATP (10 mumol/l). Whereas the evoked [3H]-noradrenaline overflow was unaffected by suramin and DIDS, cibacron blue and alpha,beta-methylene-ATP caused a small and transient increase. Cibacron blue at 30 mumol/l failed to antagonize the inhibition of evoked [3H]-noradrenaline overflow that adenosine produced in the absence of DPCPX. Basal [3H]-noradrenaline overflow was enhanced by cibacron blue, not changed by alpha,beta-methylene-ATP and DIDS, and decreased by suramin. The results show that exogenous ATP inhibits sympathetic neurotransmission in the rat iris via A1 and P2Y-like purinoceptors. The latter have a low apparent affinity for cibacron blue and probably are blocked by alpha,beta-methylene-ATP. Under the present conditions, endogenous purines exert a tonic inhibition not only via A1- but also via these P2Y-receptors.